The use of HIV-antiretroviral therapy has allowed HIV-infected individuals to live longer but has also increased the prevalence of HIV-associated neurocognitive disorders (HAND) in the U.S. and worldwide. One of the hallmarks of HAND is the infiltration of immune cells and viral replication in the brain. This process occurs as a resul of blood-brain barrier (BBB) hyperpermeability, which regulates immune cell infiltration and is traditionally low in healthy patients. Our group has been investigating changes in BBB integrity and brain endothelial remodeling as an indicator of cerebral vasculature health and a potential early biomarker for HAND. We have found that brain endothelial cells (EC) release extracellular microvesicles (eMVs) in response to HIV proteins as well as drugs of abuse. We hypothesize that HIV virotoxins, HIV infected monocytes alone or in combination with psychostimulant insult will induce a time and content-specific release of brain EC-derived eMVs, which will provide a means of identifying and monitoring of cerebrovascular health during HIV infection. The discovery of eMV production from brain ECs represents a paradigm shift in our understanding of BBB biology. The reason being that we must now consider what the implications and consequences of proteins, such as tight junction proteins, released in eMVs mean for BBB dysfunction, compensation and repair. To this end, we needed to modernize and innovate an entirely new BBB model platform in order to answer questions related to how HIV related insult affects eMV biogenesis, their temporal kinetics and eMV composition, while applying physiologically relevant conditions that would allow mechanotransduction to occur (i.e., via shear stress). Therefore, we will utilize advanced microfluidic chip technology in combination with our expertise in culturing primary cells derived from human brain tissue to generate stable quad co-cultures of endothelial cells, neurons, pericytes and astrocytes in a CNS-like compartmentalized environment. Our model recreates the BBB cyto- architecture of the neurovascular unit (NVU) and the fluidic forces that ECs experience in vivo, while allowing for real-time imaging of immune cell infiltration into the CNS-like or neuronal-astrocytic compartment. To our knowledge, we are the first to propose the use of an advanced BBB microfluidic model to study the effects of HIV proteins, HIV-infected monocytes combined with drugs of abuse on the BBB. Together the aims proposed in this study will provide insight into a novel BBB remodeling phenomenon following insult by HIV and drugs of abuse which could lead to biomarker discovery and identification of new pathologic indicators. Furthermore, this revolutionary NVU-chip has broad applicability beyond the studies proposed here; for instance screening HIV antiviral CNS penetrability, evaluation of BBB protective therapies and adjunct/alternate approaches for treating HIV in the CNS.